Inexpensive bioprinting on a microscope using liquid crystal displays and visible light

Abstract

Patterned photocrosslinking has several uses in the biofabrication of microstructurally complex tissue constructs, through both photolithography of scaffolds and photoconjugation of cell adhesive and instructive moieties. Often the polymers used are modified by methacrylation while photoactivation requires ultraviolet light. In contrast, this study aimed to design, build and evaluate a low-cost platform to place photocrosslink patterns into unmodified collagen and gelatin hydrogels using visible light and ruthenium-mediated tyrosine crosslinking in a way compatible with cell culture and inverted microscopes commonly used in biological laboratories. A photoprinting module was constructed above an inverted microscope sample stage to be confocal with the imaging system. The module consists of a blue light emitting diode array, light pipe, diffuser, microelectronically controlled liquid crystal display as photomask, and focusing objective. Resulting Ruthenium-mediated photocrosslink patterns were visible in unmodified collagen and gelatin hydrogels due to altered local polymer network density and optical contrast. Green fluorescent protein was conjugated in patterns to both gelatin and collagen gels, dependent on light exposure, intensity, and polymer network density. Pattern resolution varied from 2.0 ± 0.5 μm to 102 ± 33 μm (mean ± standard deviation) dependent on the focusing objective magnification and the pattern used (display pixel versus diode element). Further, photocrosslink patterns placed in collagen hydrogels and incubated without rinsing in serum-containing media swelled over 20–48 h, breaking the collagen network and forming ∼50 μm diameter holes. Fibroblasts cultured in photopatterned collagen hydrogels aligned and moved on and around crosslinked regions, consistent with durotaxis and contact guidance. The platform for photocrosslinking described in this study will impact several research fields, notably bioprinting of microstructurally and mechanically complex tissue constructs.